Background & Aims NADPH oxidase (NOX) generates reactive oxygen species (ROS) in hepatic stellate cells (HSCs) during liver fibrosis. In response to fibrogenic agonists, such as angiotensin II (Ang II), the NOX1 components form an active complex including Rac1. Superoxide dismutase 1 (SOD1) interacts with the NOX-Rac1 complex to stimulate NOX activity. NOX4 is also induced in activated HSCs/myofibroblast by increased gene expression. Here, we investigate the role of an enhanced activity SOD1 G37R mutation (SODmu) and the effects of GKT137831, a dual NOX1/4 inhibitor, on HSCs and liver fibrosis. Methods To induce liver fibrosis, wild-type (WT) and SOD1mu mice were treated with carbon tetrachloride (CCl4) or bile duct ligation (BDL). Then, to address the role of NOX-SOD1-mediated ROS production in HSC activation and liver fibrosis, mice were treated with a NOX1/4 inhibitor. Fibrosis and ROS generation was assessed by histology and measurement of TBARS and NOX related genes. Primary cultured HSCs isolated from WT, SODmu, and NOX1 knock-out (KO) mice were assessed for ROS production, Rac1 activity, and NOX gene expression. Results Liver fibrosis was increased in SOD1mu mice, and ROS production and Rac1 activity were increased in SOD1mu HSCs. The NOX1/4 inhibitor GKT137831 attenuated liver fibrosis and ROS production in both SOD1mu and WT mice as well as mRNA expression of fibrotic and NOX genes. Treatment with GKT137831 suppressed ROS production and NOX and fibrotic gene expression, but not Rac1 activity, in SOD1mut and WT HSCs. Both Ang II and TGFb upregulated NOX4, but AngII required NOX1. Conclusions SOD1mu induces excessive NOX1 activation through Rac1 in HSCs, causing enhanced NOX4 upregulation, ROS generation, and liver fibrosis. Treatment targeting NOX1/4 may be a new therapy for liver fibrosis.

We describe the design, synthesis, and optimization of first-in-class series of inhibitors of NADPH oxidase isoform 4 (Nox4), an enzyme implicated in several pathologies, in particular idiopathic pulmonary fibrosis, a life-threatening and orphan disease. Initially, several moderately potent pyrazolopyridine dione derivatives were found during a high-throughput screening campaign. SAR investigation around the pyrazolopyridine dione core led to the discovery of several double-digit nanomolar inhibitors in cell free assays of reactive oxygen species (ROS) production, showing high potency on Nox4 and Nox1. The compounds have little affinity for Nox2 isoform and are selective for Nox4/1 isoforms. The specificity of these compounds was confirmed in an extensive in vitro pharmacological profile, as well as in a counterscreening assay for potential ROS scavenging. Concomitant benefits are good oral bioavailability and high plasma concentrations in vivo, allowing further clinical trials for the potential treatment of fibrotic diseases, cancers, and cardiovascular and metabolic diseases.

The photophysical properties of a series of helicene cations in various solvents have been investigated using stationary and time-resolved spectroscopy. These compounds fluoresce in the near infrared region with a quantum yield ranging between 2 and 20% and a lifetime between 1 and 12 ns, depending of the solvent. No clear solvent dependence could be recognized except for a decrease of fluorescence quantum yield and lifetime with increasing hydrogen-bond donating ability of the solvent. In water, the helicene cations undergo aggregation. This effect manifests itself by the presence of a slow fluorescence decay component, whose amplitude increases with dye concentration, and by a much slower decay of the polarization anisotropy in water compared to an organic solvent of similar viscosity. However, aggregation has essentially no effect on the stationary fluorescence spectrum, whereas relatively small changes can be seen in the absorption spectrum. Analysis of the dependence of aggregation on the dye concentration reveals that the aggregates are mostly dimers and that the aggregation constant is substantially larger for hetero- than homochiral dimers.

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Addition of an enantiopure sulfoxide auxiliary is the key to the resolution of highly stable [4]helicenium dyes. This transformation allows a facile chromatographic separation of the diastereomers (the structure of one such diastereomer is shown; S yellow, O red, N blue). Then an unprecedented Pummerer‐like CC bond fragmentation takes place to release the enantiopure cations.